Molecular and Functional Characterization of Choline Transporter-Like Proteins in Esophageal Cancer Cells and Potential Therapeutic Targets

In this study, we examined the molecular and functional characterization of choline uptake in the human esophageal cancer cells. In addition, we examined the influence of various drugs on the transport of [3H]choline, and explored the possible correlation between the inhibition of choline uptake and apoptotic cell death. We found that both choline transporter-like protein 1 (CTL1) and CTL2 mRNAs and proteins were highly expressed in esophageal cancer cell lines (KYSE series). CTL1 and CTL2 were located in the plasma membrane and mitochondria, respectively. Choline uptake was saturable and mediated by a single transport system, which is both Na+-independent and pH-dependent. Choline uptake and cell viability were inhibited by various cationic drugs. Furthermore, a correlation analysis of the potencies of 47 drugs for the inhibition of choline uptake and cell viability showed a strong correlation. Choline uptake inhibitors and choline deficiency each inhibited cell viability and increased caspase-3/7 activity. We conclude that extracellular choline is mainly transported via a CTL1. The functional inhibition of CTL1 by cationic drugs could promote apoptotic cell death. Furthermore, CTL2 may be involved in choline uptake in mitochondria, which is the rate-limiting step in S-adenosylmethionine (SAM) synthesis and DNA methylation. Identification of this CTL1- and CTL2-mediated choline transport system provides a potential new target for esophageal cancer therapy.

Novel non-apoptotic cell death: ferroptosis / 영남의대학술지

Ferroptosis is a newly recognized type of cell death that results from iron-dependent lipid peroxidation and is different from other types of cell death, such as apoptosis, necrosis, and autophagic cell death. This type of cell death is characterized by mitochondrial shrinkage with an increased mitochondrial membrane density and outer mitochondrial membrane rupture. Ferroptosis can be induced by a loss of activity of system Xc− and the inhibition of glutathione peroxidase 4, followed by the accumulation of lipid reactive oxygen species (ROS). In addition, inactivation of the mevalonate and transsulfuration pathways is involved in the induction of ferroptosis. Moreover, nicotinamide adenine dinucleotide phosphate oxidase and p53 promote ferroptosis by increasing ROS production, while heat shock protein beta-1 and nuclear factor erythroid 2-related factor 2 inhibit ferroptosis by reducing iron uptake. This article outlines the molecular mechanisms and signaling pathways of ferroptosis regulation, and explains the roles of ferroptosis in human disease.

Novel non-apoptotic cell death: ferroptosis / 영남의대학술지

Ferroptosis is a newly recognized type of cell death that results from iron-dependent lipid peroxidation and is different from other types of cell death, such as apoptosis, necrosis, and autophagic cell death. This type of cell death is characterized by mitochondrial shrinkage with an increased mitochondrial membrane density and outer mitochondrial membrane rupture. Ferroptosis can be induced by a loss of activity of system Xc− and the inhibition of glutathione peroxidase 4, followed by the accumulation of lipid reactive oxygen species (ROS). In addition, inactivation of the mevalonate and transsulfuration pathways is involved in the induction of ferroptosis. Moreover, nicotinamide adenine dinucleotide phosphate oxidase and p53 promote ferroptosis by increasing ROS production, while heat shock protein beta-1 and nuclear factor erythroid 2-related factor 2 inhibit ferroptosis by reducing iron uptake. This article outlines the molecular mechanisms and signaling pathways of ferroptosis regulation, and explains the roles of ferroptosis in human disease.

Effects of LED irradiation on the expression of apoptosis-related molecules in human SH-SY5Y neuroblastoma cells

To verify the inhibitory or protective effects of light-emitting diode(LED) irradiation on apoptotic cell death induced by CoCl2, human SH-SY5Y cells were treated with CoCl2 and LED were used to irradiate the cells. In the cell viability assay, cells were died slowly from 50 micrometer to 250 micrometer and about 50% of cells died after 12 hours at 400 micrometer of CoCl2. The Diff-Quik staining revealed that cells showed condensation of DNA and blebbing of the cell membrane. The DNA fragmentation assay revealed the DNA fragmentation, which is another apoptosis marker, occurred in cells treated with 400 micrometer CoCl2 for 16 hours. In the western blot for HIF-1 alpha, HIF-1 alpha was expressed after 3 hours from induction and peaked maximally at 16 hours. In the cell viability assay of the effects of LED irradiation (at 590 nm for 1 hour 20 minutes), the cells showed more proliferation (about 20%) than the control group. The RPA assay of various apoptosis-related molecules showed that pro-apoptosis molecules such as Bax, Bak, and Bid were upregulated in the CoCl2 treatment group. This means that the apoptotic cell population was increased. However there was some significant changes in LED irradiated cells. In the CoCl2-treated LED irradiation group, those molecules were down-regulated more than in the only CoCl2-treated group. These results have shown that CoCl2 may induce apoptotic cell death in human SH-SY5Y neuroblastoma cells. And LED irradiation has a positive effect on apoptotic cells by down-regulation of pro-apoptotic molecules.

Effect of Acupuncture on Apoptosis and Protein Caspase-9 Expression in Hippocampus of Cerebral Ischemia Rats / 中国中医药信息杂志

Objective To observe the effect of acupuncture on apoptosis and Caspase-9 expression in hippocampus of cerebral ischemia (CI) rats. Methods Thirty male SD rats were randomized into control, CI and CI + acupuncture group, with 10 cases in each group. CI model was established by occlusion of the unilateral middle cerebral artery. "Baihui" (GV20) and "Shuigou" (GV26) were acupunctured once daily for 7 days. The apoptosis and expression of protein Caspase-9 in hippocampus was displayed by HE staining and immunohistochemical method. Results In comparison with the normal control group, Caspase-9 immune reaction (IR) positive cells in the hippocampus of CI group increased significantly (P

Application of anti-ICAM McAb in rejection after liver transplantation and hepatocellular apoptosis in rats / 中华肝胆外科杂志

Objective To compare the effects among 1A29, CsA and their combined use on rejection after liver transplantation and hepatocellular apoptosis in rats. Methods The stable rat model of liver transplantation was established. Rats of the same strain were employed as the control group. The effects of 1A29, CsA and their combined use on rejection after liver transplantation were determined in both groups. Meanwhile, the hepatocellular apoptosis was recorded and evaluated. Results No rejection was found between the rats of the same strain (SDSD), while different levels of rejection was seen between the rats of different strains (WistarSD). Biochemical test showed a significant increase in levels of enzyme spectrum and bile in the blood. An apparent pathological change due to rejection was also observed. CsA of optimal dose (10mg/kg) effectively suppressed the rejection while 1A29 of optimal dose did not. When used alone, CsA of sub-optimal dose had no effect on rejection. On the contrary, the combined use of CsA of sub-optimal dose and 1A29 of optimal dose could significantly inhibit the rejection. There was a marked increase of the hepatocellular apoptosis in the liver graft rejected by the recipient. However, the level of hepatocellular apoptosis showed no remarkable change in the liver graft without rejection due to use of immunosuppressors. Conclusions Hepatocellular apoptosis is closely related to rejection of the liver graft. The apoptosis is not significantly correlated to application of 1A29. The use of 1A29 can result in significant decrease in dose of CsA. Using 1A29 alone can not effectively inhibit the rejection after liver transplantation.

Effects of ginsenoside on cellular model of Alzheimer disease induced by protein phosphatase inhibitor okadaic acid / 中国药理学通报

Aim To investigate the effects of ginsenoside(GS) on phosphorylation of tau protein,microtubule,cellular apoptosis and its related factors in cellular model of Alzheimer disease(AD) induced by the protein phosphatase 1 and 2A inhibitor okadaic acid(OA).Methods The human neuroblastoma cell line SK-N-SH cells were cultured with GS for 24 h,the culture medium was changed,and then incubated with OA 10 nmol?L-1 for 6 h.The changes of cell morphology were observed by inverted microscope.The laser confocal microscopy was used to observe the microtubule changes.Western blot was applied to determine the expression of phosphorylation of tau protein,and apoptosis-regulating factors Bcl-2,Bax and Caspase-3.The changes of apoptotic cells were observed by TUNEL method.Results The normal SK-N-SH cells spread well.OA-treated cells showed that the cell axons and the microtubules were broken and decreased under the inverted microscope and laser confocal microscope.Preincubation of GS demonstrated the significantly protective effects against the morphologic damage induced by OA.In OA-treated group,the phosphorylation of tau protein at Ser-199/202 and Ser-404 sites was higher than that in normal group,and the non-phosphorylation of tau protein at the same sites was lower;Incubation of GS at the dose of 50 mg?L-1 and 100 mg?L-1 with the cells decreased the phosphorylation of tau protein Ser-199/202 and Ser-404 sites.GS group at the dose of 50 mg?L-1 and 100 mg?L-1 decreased the expression of at non-phosphorylation of tau protein at the Ser202 site.The apoptotic cells were not found in normal group.The number of apoptotic cells were obviously increased.the expression of Bax and caspase-3 significantly enhanced,and Bcl-2 expression decreased in the OA-treated model group.GS significantly decreased the apoptotic cell number of nerve cells,inhibited the expression of Bax and caspase-3.Conclusion GS can protect the nerve cells from pathological change induced by OA.Maybe because it can inhibit the hyperphosphorylation of tau protein and protect the nerve cells from apoptosis,thus GS may have potential to treat Alzheimer disease.

The Effect of Nitric Oxide on Cultured Bovine Retinal Pigment Epithelial Cell

Nitric oxide (NO), a potencially toxic radical, is generally regarded as a multi-potent molecule to be implicated in a wide range of biological function. The presence of nitric oxide synthase(NOS) in the retina, the constitutive isoform in photoreceptor outer segments and and the inducible form in retinal pigment epithelial (RPE) cells, has been demonstrated. The effect of NO in retina has been studying mainly as neurotransmitter. Present study was undertaken to find the role of NO in cultured bovine retinal pigment epithelial cells. Cultured bovine retinal pigment epithelial cells were treated with various concentrations of NO generator, S-nitroso-N-acetyl-D, L-penicillamine (SNAP). The survival fractions were measured by MTT assay. The morphologic changes were observed with inverted phase contrast microscope and electron microscope. To evaluate the characteristics of cell death, cells were lysed for DNA extraction, and the agarose gel electrophoresis was done. NO brought a decrease in the survival fraction of cultured bovine retinal pigment epithelial cells as concentrations increased. At high concentrations, cells became sparse. Electron microscopic study showed destruction of nuclear membrane and chromatin condensation in 1 mM SNAP treated group. These findings were compatible with apoptotic cell death that was supported with DNA laddering pattern in agarose gel electrophoresis. NO can induce damage to retinal pigment epithelial cells, and damaged cells are destined to apoptotic cell death.